CRISPR-Cas type II-based Synthetic Biology applications in eukaryotic cells

Area of Science:

• Synthetic Biology
• Molecular Biology
• Gene Editing

Background:

• The CRISPR-Cas system is a popular tool for DNA editing and genome engineering.
• CRISPR-Cas type II system has been adapted to create RNA-based transcription factors for synthetic biology.
• These factors consist of a CRISPR RNA and a nuclease-deficient Cas9 fused to regulatory domains.

Purpose of the Study:

• To explore the application of CRISPR-Cas technology in synthetic biology for gene regulation.
• To investigate the construction and efficacy of novel CRISPR-based transcription factors.
• To compare the simplicity of CRISPR-based transcription factors with other synthetic proteins.

Main Methods:

• Utilized CRISPR RNA to target specific promoter sequences.
• Employed a catalytically inactive Cas9 (dCas9) fused to transcriptional activation or repression domains.
• Applied these constructs for gene up- and downregulation in mammalian and yeast cells.

Main Results:

• Successfully achieved upregulation and downregulation of single genes in both mammalian and yeast cells.
• Demonstrated that CRISPR-based transcription factors are simpler to construct compared to Transcription Activator-Like effectors (TALEs).

Conclusions:

• CRISPR-based transcription factors offer a promising and simpler alternative for gene regulation in synthetic biology.
• Further research is needed to prove the feasibility of complex synthetic networks using CRISPR-dCas9 technology.
• Investigating different CRISPR types is essential for advancing this technology.